Development of a surrogate liquid biopsy from the aqueous humor in retinoblastoma eyes.

摘要 视网膜母细胞瘤是一种原发于儿童眼内的癌症。肿瘤的发生开始了 通过RB1基因的突变，这是第一个被描述的肿瘤抑制基因。对肿瘤进行调查 受RB1调控的抑制子通路为研究糖尿病的遗传机制提供了前所未有的见解。 肿瘤发生，不仅是视网膜母细胞瘤，而且几乎所有人类癌症。对具体的评价 突变、癌基因和抑癌基因、肿瘤标志物和基因表达谱已经彻底改变了 照顾乳癌、肺癌、前列腺癌等癌症患者。然而，尽管视网膜母细胞瘤相关 研究启动了癌症遗传学领域，利用这一知识来个性化治疗 视网膜母细胞瘤患者一直难以捉摸，这在很大程度上是因为我们不能安全地活组织检查这个肿瘤，因为 对眼外扩散的担忧。因此，我们目前还不能将基因和基因组的变化联系起来。 肿瘤水平与临床转归的关系。识别这些基因组变化，并提供临床相关性 对于视网膜母细胞瘤患者来说是非常必要的。这项提议的目标是开发一种代理肿瘤 视网膜母细胞瘤的活组织检查使用房水，房水是眼睛不同部位的透明液体 就是肿瘤形成的地方。从2012年开始，摘除房水的做法是常规和安全的，就像 向眼玻璃体腔内注射化疗的标准部分；没有病例 眼外扩散已有报道。我们对水溶液进行评估的初步数据显示 存在肿瘤来源的无细胞DNA，全基因组测序显示了染色体的特征 证实肿瘤变化的得失。这项研究的具体目的是为了测试 以下假设：房水可作为替代肿瘤活检--更准确地说，房水 含有可被检测并与临床结果相关的肿瘤来源的遗传物质。目标1是 为了定义房水中高度重复的染色体变化，如2p增益/MYCN扩增，这是 已知发生在视网膜母细胞瘤肿瘤中。目标2是确定特定的rb1突变(S) 房水中的肿瘤形成。最后，Aim 3将评估RNA表达谱以定义 房水中的亚群，并与肿瘤复发等临床结果相关。目前， 唯一可能实现这些目标的方法是对肿瘤组织进行评估，只有当眼睛 被移除(例如，去核)。然而，通过在水中定义这些基因变化，我们将第一次 时间，能够在治疗过程中确定原位肿瘤的特征。这将使预测的发展成为可能 治疗反应的标记物，并在未来可能指导治疗。房水的使用作为一种 代用肿瘤活检具有巨大的潜力，因为它直接影响我们处理视网膜母细胞瘤的方式 最终允许为这种致盲的儿童癌症开发个性化药物。 好了！ 好了！

Abstract Retinoblastoma is a primary intraocular cancer that develops in the eyes of children. Tumorigenesis is initiated by a mutation in the RB1 gene, which was the first tumor suppressor gene described. Investigating the tumor suppressor pathway regulated by RB1 has provided unprecedented insights into the genetic mechanisms of tumorigenesis, not only for retinoblastoma, but virtually all human cancers. The evaluation of specific mutations, oncogenes and suppressors, tumor markers and gene expression profiles have revolutionized the care of breast, lung, prostate and other cancer patients. However, despite the fact that retinoblastoma-related research launched the field of cancer genetics, leveraging this knowledge to personalize the care for retinoblastoma patients has been elusive in large part because we cannot safely biopsy this tumor due to concerns of extraocular spread. Thus, we currently cannot correlate the genetic and genomic changes at the level of the tumor with clinical outcomes. Identifying these genomic changes, and providing clinical correlation is critically needed for retinoblastoma patients. The objective of this proposal is to develop a surrogate tumor biopsy for retinoblastoma using the aqueous humor, which is the clear fluid in a separate part of the eye from where the tumor forms. Starting in 2012, extractions of aqueous humor have been routinely and safely done as a standard part of the procedure to inject chemotherapy into the vitreous cavity of the eye; no cases of extraocular spread have been reported. Our preliminary data from evaluation of the aqueous revealed that tumor-derived cell-free DNA is present and whole genome sequencing demonstrates a profile of chromosomal gains and losses that corroborate changes in the tumor. The specific aims of this study are designed to test the following hypothesis: the aqueous can serve as a surrogate tumor biopsy – more precisely, the aqueous harbors tumor-derived genetic material which can be assayed and correlated with clinical outcomes. Aim 1 is to define highly recurrent chromosomal changes in the aqueous, such as 2p gain/MYCN amplification, that are known to occur in retinoblastoma tumors. Aim 2 is to identify the specific RB1 mutation(s) that underlie tumorigenesis from the aqueous humor. Finally, Aim 3 will evaluate RNA expression profiles to define subgroups in the aqueous, and correlate these with clinical outcomes such as tumor recurrence. Currently, the only way any of these Aims are possible is from evaluation of tumor tissue, which is only available if an eye is removed (e.g. enucleated). However, by defining these genetic changes in the aqueous we will, for the first time, be able to characterize tumors in situ during therapy. This will allow for the development of prognostic markers for therapeutic response and, in the future, may guide therapy. Use of the aqueous humor as a surrogate tumor biopsy holds tremendous potential, from directly impacting the way we manage retinoblastoma patients, to ultimately allowing for the development of personalized medicine for this blinding childhood cancer. ! !